Objective To develop a new type of gene vaccine against plague. Methods caf1 gene of caf Operon Encoding F1 Antigen of Y. pestis was inserted into pHSS6-mTn-3xHA/lacZ library plasmids, the recombinant plasmids were linearized with Not I and transformed into yeast cell by lithium acetate (LiAc) method to induce homologous recombination. Positive recombinants were then selected with uracil-lack medium. Results These recombinants were confirmed by colony PCR to have the target gene fragments. Conclusion The present study provided a platform for constructing a novel expression system for expressing Y. pestis F1 Antigen via homologous recombination in Saccharomyces cerevisiae, which may contribute to the genetic prevention of plague through digestive-tract-route (DTR).